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GENOME-WIDE COPY NUMBER PROFILING OF RENAL CELL TUMORS USING ARRAY-CGH
Vol. 179, No. 4, Supplement, Sunday, May 18, 2008
compared to clear cell or papillary tumors. It appears that most RCC subtypes possess multiple and redundant mechanisms that thwart the antitumoral immune response. Source of Funding: None
379 OVEREXPRESSION OF XIAP EXPRESSION IN RENAL CELL CARCINOMA PREDICTS WORSE PROGNOSIS Yoichi Mizutani*, Hiroyuki Nakanishi, Takumi Shiraishi, Terukazu Nakamura, Kazuya Mikami, Natsuki Takaha, Koji Okihara, Osamu Ukimura, Akihiro Kawauchi, Tsuneharu Miki. Kyoto, Japan. INTRODUCTION AND OBJECTIVE: X-linked inhibitor of apoptosis protein ( XIAP ) is the most potent caspase-inhibitory IAP family member which blocks apoptosis. However, little is known about WKHFOLQLFDOVLJQL¿FDQFHRI;,$3LQYDULRXVFDQFHUVLQFOXGLQJUHQDOFHOO carcinoma ( RCC ). This study examined XIAP expression in 109 normal kidneys and RCCs. METHODS: The level of XIAP expression was determined E\:HVWHUQEORWDQDO\VLVXVLQJQRQ¿[HGIUHVKIUR]HQWLVVXHVRI5&&V and normal kidneys. RESULTS: The mean level of XIAP expression was higher in RCC compared to autologous normal kidney. In Stage I/II RCC, the mean XIAP expression level was almost identical to that in normal kidney, whereas XIAP expression in Stage III/IV was 2.5-fold higher. The level of XIAP expression correlated with the grade of RCC. Patients with RCC with low XIAP expression had a longer postoperative diseaseVSHFL¿FVXUYLYDOWKDQWKRVHZLWKKLJKH[SUHVVLRQLQWKH\HDUIROORZXS Treatment of RCC cells with XIAP antisense oligonucleotide enhanced Fas-mediated and tumor necrosis factor-related apoptosis-inducing ligand ( TRAIL )-mediated apoptosis. CONCLUSIONS: The present study is the first study to demonstrate that XIAP expression was enhanced in RCC, and that high XIAP expression in RCC predicted worse prognosis. In addition, ;,$3DQWLVHQVHROLJRQXFOHRWLGHVHQVLWL]HG5&&WR)DV75$,/LQGXFHG apoptosis. These results suggest that XIAP expression in RCC may be used as a prognostic parameter, and that downregulation of XIAP expression in RCC may potentiate immunotherapy. Source of Funding: None
380 GENOME-WIDE COPY NUMBER PROFILING OF RENAL CELL TUMORS USING ARRAY-CGH Jimsgene Sanjmyatav, Carsten Schwaenen, Sven Wessendorf, Jorg Schubert, Kerstin Junker*. Jena, Germany, and Ulm, Germany. INTRODUCTION AND OBJECTIVE: The TNM-staging system has been shown to be the most accurate tool to predict patient outcome in RCC. The aim of this study was to investigate a possible relation between clinical TNM staging and genetic properties of tumors using high-resolution array-CGH, and to identify novel chromosomal abnormalities which could have prognostic value. METHODS: We conducted genome-wide copy number SUR¿OHV LQ SULPDU\ UHQDO FHOO WXPRUV LQFOXGLQJ FOHDU FHOO papillary, 13 chromophobe RCC and 13 oncocytomas by array-CGH with a median resolution of 1-1,5MB. The bioinformatic evaluation of array-data was performed by the software XLminer. RESULTS: Although tumors with advanced TNM-status DFFXPXODWHGPRUHFKURPRVRPDODOWHUDWLRQVQRVLJQL¿FDQWUHODWLRQVKLS between copy number changes and tumor stages of RCC could be found. For differentiation of RCC entities XLminer generated rules allowing to identify tumors by array patterns.Thirty-four percent of clear cell, 30% of papillary, 46% of chromophobe RCC and 30% of oncocytomas revealed DJDLQRIWKHORFXVTRQHRIWKHPRVWIUHTXHQWO\DPSOL¿HGUHJLRQV within the human genome where six oncogenes (CCND1, FGF4, FGF3, EMS1, GARP, and PAK1) are located. FISH using 2 BAC-clones on this UHJLRQFRQ¿UPHGWKH¿QGLQJVRIDUUD\&*+$FFRUGLQJWRWKH;/PLQHU evaluation, the gain of 11q13.4 was not associated with metastasis LQ 5&& )XUWKHUPRUH ZH ZHUH DEOH WR GH¿QH VSHFL¿F UHJLRQV RQ chromosomes 8, 9 and 14 for differentiation between metastatic and non-metastatic tumors.
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CONCLUSIONS: Our results suggest that chromosomal alterations in RCC are independent of the TNM status of tumors. It is SRVVLEOHWRGH¿QHVSHFL¿FFKURPRVRPDOUHJLRQVZKLFKDUHDVVRFLDWHG with metastatic disease. To predict comprehensively biological behavior of tumors further studies regarding chromosomal alterations, gene expression and protein expression are needed. Source of Funding: Deutsche Krebshilfe
381 DECREASED EXPRESSION OF TRANSCRIPTION FACTOR 2 (TCF-2) INDICATES POOR PROGNOSIS IN METASTATIC RENAL CELL CARCINOMA – VALIDATION OF MICROARRAY DATA Alexander Buchner*, Mirna Castro, Anja Hennig, Gerald Assmann, Alfons Hofstetter, Christian G Stief, Wolfgang Zimmermann. Munich, Germany. INTRODUCTION AND OBJECTIVE: The survival of patients with metastatic renal cell carcinoma (mRCC) varies remarkably, even among patients with similar tumor stage. Performing gene expression SUR¿OLQJ ZLWK ODVHU PLFURGLVVHFWLRQ DQG ROLJRQXFOHRWLGH PLFURDUUD\V ZH KDYH LGHQWL¿HG7&) DV D SDUW RI D WKUHHJHQH VLJQDWXUH WKDW LV DQLQGHSHQGHQWSURJQRVWLFIDFWRULQP5&&,QWKLVVWXG\ZHDQDO\]HG the expression of TCF-2 on RNA and protein level and its impact on patients’ survival. METHODS: Gene expression profiling was done on 28 samples of clear-cell RCC metastases (patients’ follow-up time up to 116 months) using laser microdissection and oligonucleotide microarrays (Affymetrix GeneChip® HG U133 Plus 2.0). A prognosis-associated gene VLJQDWXUHZDVLGHQWL¿HGXVLQJWKHVHPLVXSHUYLVHGSULQFLSDOFRPSRQHQWV DQDO\VLV PHWKRG WKDW XWLOL]HV WKH LQGLYLGXDO VXUYLYDO GDWD %DLU DQG Tibshirani 2004). Validation was performed with samples of normal renal tissue (n=6), RCC primary tumor (n=57) and RCC metastases (n=59) WKDW ZHUH DQDO\]HG E\ TXDQWLWDWLYH 573&5 ,PPXQRKLVWRFKHPLVWU\ was done in a subset of the tissue samples using a polyclonal antibody against TCF-2. RESULTS: Analysis of expression data revealed a threegene signature that discriminated well between two patient groups with different outcome (p<0.05). TCF-2 was part of this gene signature DQG ZDV DQDO\]HG LQ GHWDLO 7KH 573&5 GDWD VKRZHG D VLJQL¿FDQW decrease in TCF-2 expression correlated with malignant transformation QRUPDOUHQDOWLVVXH!SULPDU\WXPRU!PHWDVWDVLV S 3DWLHQWV ZLWKKLJK7&)H[SUHVVLRQLQSULPDU\WXPRUKDGDVLJQL¿FDQWO\EHWWHU prognosis (p<0.05). TCF-2 immunohistochemistry showed a nuclear VWDLQLQJFRQ¿QHGWRWKHWXPRUFHOOVERWKRIWKHSULPDU\WXPRUVDQGRI the metastases. Tubuli of normal renal tissue showed strong TCF-2 reaction, while stroma cells were negative. CONCLUSIONS: The expression of TCF-2 in tumor cells is DUHOHYDQWSURJQRVWLFIDFWRULQP5&&ZLWKDVLJQL¿FDQWGHFUHDVHRIWKH RNA expression level correlated with malignant transformation. Patients ZLWKKLJK7&)H[SUHVVLRQKDYHDVLJQL¿FDQWO\EHWWHUSURJQRVLV7KHVH UHVXOWVFRQ¿UPWKHPLFURDUUD\¿QGLQJVIURPODVHUPLFURGLVVHFWHG5&& tissue. Measurement of the TCF-2 RNA level could help to stratify mRCC patients for optimal therapy in relation to their prognosis. Source of Funding: German Cancer Aid.
382 THE ROLE OF LYMPHOVASCULAR INVASION IN RENAL CELL CARCINOMA AS A PROGNOSTIC MARKER OF SURVIVAL Matthew D Katz*, Maria F Serrano, Yan Yan, Peter A Humphrey, Adam S Kibel. Saint Louis, MO. INTRODUCTION AND OBJECTIVE: The most common pathologic correlates of renal carcinoma (RCC) aggressiveness are Fuhrman grade, TNM stage, and histologic subtype. Lymphovascular invasion (LVI) correlates with adverse outcomes in numerous malignancies. However, its role in predicting outcomes in RCC is unclear. 3ULRUZRUNKDVIRXQGDQDVVRFLDWLRQZLWKFDQFHUVSHFL¿FVXUYLYDO+HUHLQ we evaluated what effect LVI within the primary tumor has on metastasis IUHHVXUYLYDO 0)6 FDQFHUVSHFL¿FVXUYLYDO &66 DQGRYHUDOOVXUYLYDO (OS) in patients with RCC treated with surgical excision. 0(7+2'6 :HLGHQWL¿HG SDWLHQWVZKRXQGHUZHQWHLWKHU a partial or radical nephrectomy for RCC at our institution from 1989 to
compared to clear cell or papillary tumors. It appears that most RCC subtypes possess multiple and redundant mechanisms that thwart the antitumoral immune response. Source of Funding: None
379 OVEREXPRESSION OF XIAP EXPRESSION IN RENAL CELL CARCINOMA PREDICTS WORSE PROGNOSIS Yoichi Mizutani*, Hiroyuki Nakanishi, Takumi Shiraishi, Terukazu Nakamura, Kazuya Mikami, Natsuki Takaha, Koji Okihara, Osamu Ukimura, Akihiro Kawauchi, Tsuneharu Miki. Kyoto, Japan. INTRODUCTION AND OBJECTIVE: X-linked inhibitor of apoptosis protein ( XIAP ) is the most potent caspase-inhibitory IAP family member which blocks apoptosis. However, little is known about WKHFOLQLFDOVLJQL¿FDQFHRI;,$3LQYDULRXVFDQFHUVLQFOXGLQJUHQDOFHOO carcinoma ( RCC ). This study examined XIAP expression in 109 normal kidneys and RCCs. METHODS: The level of XIAP expression was determined E\:HVWHUQEORWDQDO\VLVXVLQJQRQ¿[HGIUHVKIUR]HQWLVVXHVRI5&&V and normal kidneys. RESULTS: The mean level of XIAP expression was higher in RCC compared to autologous normal kidney. In Stage I/II RCC, the mean XIAP expression level was almost identical to that in normal kidney, whereas XIAP expression in Stage III/IV was 2.5-fold higher. The level of XIAP expression correlated with the grade of RCC. Patients with RCC with low XIAP expression had a longer postoperative diseaseVSHFL¿FVXUYLYDOWKDQWKRVHZLWKKLJKH[SUHVVLRQLQWKH\HDUIROORZXS Treatment of RCC cells with XIAP antisense oligonucleotide enhanced Fas-mediated and tumor necrosis factor-related apoptosis-inducing ligand ( TRAIL )-mediated apoptosis. CONCLUSIONS: The present study is the first study to demonstrate that XIAP expression was enhanced in RCC, and that high XIAP expression in RCC predicted worse prognosis. In addition, ;,$3DQWLVHQVHROLJRQXFOHRWLGHVHQVLWL]HG5&&WR)DV75$,/LQGXFHG apoptosis. These results suggest that XIAP expression in RCC may be used as a prognostic parameter, and that downregulation of XIAP expression in RCC may potentiate immunotherapy. Source of Funding: None
380 GENOME-WIDE COPY NUMBER PROFILING OF RENAL CELL TUMORS USING ARRAY-CGH Jimsgene Sanjmyatav, Carsten Schwaenen, Sven Wessendorf, Jorg Schubert, Kerstin Junker*. Jena, Germany, and Ulm, Germany. INTRODUCTION AND OBJECTIVE: The TNM-staging system has been shown to be the most accurate tool to predict patient outcome in RCC. The aim of this study was to investigate a possible relation between clinical TNM staging and genetic properties of tumors using high-resolution array-CGH, and to identify novel chromosomal abnormalities which could have prognostic value. METHODS: We conducted genome-wide copy number SUR¿OHV LQ SULPDU\ UHQDO FHOO WXPRUV LQFOXGLQJ FOHDU FHOO papillary, 13 chromophobe RCC and 13 oncocytomas by array-CGH with a median resolution of 1-1,5MB. The bioinformatic evaluation of array-data was performed by the software XLminer. RESULTS: Although tumors with advanced TNM-status DFFXPXODWHGPRUHFKURPRVRPDODOWHUDWLRQVQRVLJQL¿FDQWUHODWLRQVKLS between copy number changes and tumor stages of RCC could be found. For differentiation of RCC entities XLminer generated rules allowing to identify tumors by array patterns.Thirty-four percent of clear cell, 30% of papillary, 46% of chromophobe RCC and 30% of oncocytomas revealed DJDLQRIWKHORFXVTRQHRIWKHPRVWIUHTXHQWO\DPSOL¿HGUHJLRQV within the human genome where six oncogenes (CCND1, FGF4, FGF3, EMS1, GARP, and PAK1) are located. FISH using 2 BAC-clones on this UHJLRQFRQ¿UPHGWKH¿QGLQJVRIDUUD\&*+$FFRUGLQJWRWKH;/PLQHU evaluation, the gain of 11q13.4 was not associated with metastasis LQ 5&& )XUWKHUPRUH ZH ZHUH DEOH WR GH¿QH VSHFL¿F UHJLRQV RQ chromosomes 8, 9 and 14 for differentiation between metastatic and non-metastatic tumors.
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135
CONCLUSIONS: Our results suggest that chromosomal alterations in RCC are independent of the TNM status of tumors. It is SRVVLEOHWRGH¿QHVSHFL¿FFKURPRVRPDOUHJLRQVZKLFKDUHDVVRFLDWHG with metastatic disease. To predict comprehensively biological behavior of tumors further studies regarding chromosomal alterations, gene expression and protein expression are needed. Source of Funding: Deutsche Krebshilfe
381 DECREASED EXPRESSION OF TRANSCRIPTION FACTOR 2 (TCF-2) INDICATES POOR PROGNOSIS IN METASTATIC RENAL CELL CARCINOMA – VALIDATION OF MICROARRAY DATA Alexander Buchner*, Mirna Castro, Anja Hennig, Gerald Assmann, Alfons Hofstetter, Christian G Stief, Wolfgang Zimmermann. Munich, Germany. INTRODUCTION AND OBJECTIVE: The survival of patients with metastatic renal cell carcinoma (mRCC) varies remarkably, even among patients with similar tumor stage. Performing gene expression SUR¿OLQJ ZLWK ODVHU PLFURGLVVHFWLRQ DQG ROLJRQXFOHRWLGH PLFURDUUD\V ZH KDYH LGHQWL¿HG7&) DV D SDUW RI D WKUHHJHQH VLJQDWXUH WKDW LV DQLQGHSHQGHQWSURJQRVWLFIDFWRULQP5&&,QWKLVVWXG\ZHDQDO\]HG the expression of TCF-2 on RNA and protein level and its impact on patients’ survival. METHODS: Gene expression profiling was done on 28 samples of clear-cell RCC metastases (patients’ follow-up time up to 116 months) using laser microdissection and oligonucleotide microarrays (Affymetrix GeneChip® HG U133 Plus 2.0). A prognosis-associated gene VLJQDWXUHZDVLGHQWL¿HGXVLQJWKHVHPLVXSHUYLVHGSULQFLSDOFRPSRQHQWV DQDO\VLV PHWKRG WKDW XWLOL]HV WKH LQGLYLGXDO VXUYLYDO GDWD %DLU DQG Tibshirani 2004). Validation was performed with samples of normal renal tissue (n=6), RCC primary tumor (n=57) and RCC metastases (n=59) WKDW ZHUH DQDO\]HG E\ TXDQWLWDWLYH 573&5 ,PPXQRKLVWRFKHPLVWU\ was done in a subset of the tissue samples using a polyclonal antibody against TCF-2. RESULTS: Analysis of expression data revealed a threegene signature that discriminated well between two patient groups with different outcome (p<0.05). TCF-2 was part of this gene signature DQG ZDV DQDO\]HG LQ GHWDLO 7KH 573&5 GDWD VKRZHG D VLJQL¿FDQW decrease in TCF-2 expression correlated with malignant transformation QRUPDOUHQDOWLVVXH!SULPDU\WXPRU!PHWDVWDVLV S 3DWLHQWV ZLWKKLJK7&)H[SUHVVLRQLQSULPDU\WXPRUKDGDVLJQL¿FDQWO\EHWWHU prognosis (p<0.05). TCF-2 immunohistochemistry showed a nuclear VWDLQLQJFRQ¿QHGWRWKHWXPRUFHOOVERWKRIWKHSULPDU\WXPRUVDQGRI the metastases. Tubuli of normal renal tissue showed strong TCF-2 reaction, while stroma cells were negative. CONCLUSIONS: The expression of TCF-2 in tumor cells is DUHOHYDQWSURJQRVWLFIDFWRULQP5&&ZLWKDVLJQL¿FDQWGHFUHDVHRIWKH RNA expression level correlated with malignant transformation. Patients ZLWKKLJK7&)H[SUHVVLRQKDYHDVLJQL¿FDQWO\EHWWHUSURJQRVLV7KHVH UHVXOWVFRQ¿UPWKHPLFURDUUD\¿QGLQJVIURPODVHUPLFURGLVVHFWHG5&& tissue. Measurement of the TCF-2 RNA level could help to stratify mRCC patients for optimal therapy in relation to their prognosis. Source of Funding: German Cancer Aid.
382 THE ROLE OF LYMPHOVASCULAR INVASION IN RENAL CELL CARCINOMA AS A PROGNOSTIC MARKER OF SURVIVAL Matthew D Katz*, Maria F Serrano, Yan Yan, Peter A Humphrey, Adam S Kibel. Saint Louis, MO. INTRODUCTION AND OBJECTIVE: The most common pathologic correlates of renal carcinoma (RCC) aggressiveness are Fuhrman grade, TNM stage, and histologic subtype. Lymphovascular invasion (LVI) correlates with adverse outcomes in numerous malignancies. However, its role in predicting outcomes in RCC is unclear. 3ULRUZRUNKDVIRXQGDQDVVRFLDWLRQZLWKFDQFHUVSHFL¿FVXUYLYDO+HUHLQ we evaluated what effect LVI within the primary tumor has on metastasis IUHHVXUYLYDO 0)6 FDQFHUVSHFL¿FVXUYLYDO &66 DQGRYHUDOOVXUYLYDO (OS) in patients with RCC treated with surgical excision. 0(7+2'6 :HLGHQWL¿HG SDWLHQWVZKRXQGHUZHQWHLWKHU a partial or radical nephrectomy for RCC at our institution from 1989 to